Inter-residual Hydrogen Bonding in Carbohydrates Unraveled by NMR Spectroscopy and Molecular Dynamics Simulations.

Carbohydrates, also known as glycans in biological systems, are omnipresent in nature where they as glycoconjugates occur as oligo- and polysaccharides linked to lipids and proteins. Their three-dimensional structure is defined by two or three torsion angles at each glycosidic linkage. In addition, transglycosidic hydrogen bonding between sugar residues may be important. Herein we investigate the presence of these inter-residue interactions by NMR spectroscopy in D2 O/[D6 ]DMSO (70:30) or D2 O and by molecular dynamics (MD) simulations with explicit water as solvent for disaccharides with structural elements α-d-Manp-(1→2)-d-Manp, ß-d-GlcpNAc-(1→2)-d-Manp, and α-d-Glcp-(1→4)-ß-d-Glcp, all of which have been suggested to exhibit inter-residue hydrogen bonding. For the disaccharide ß-d-GlcpNAc-(1→2)-ß-d-Manp-OMe, the large extent of O5'⋅⋅⋅HO3 hydrogen bonding as seen from the MD simulation is implicitly supported by the 1 H NMR chemical shift and 3 JHO3,H3 value of the hydroxy proton. In the case of α-d-Glcp-(1→4)-ß-d-Glcp-OMe, the existence of a transglycosidic hydrogen bond O2'⋅⋅⋅HO3 was proven by the presence of a cross-peak in 1 H,13 C HSQC-TOCSY experiments as a result of direct TOCSY transfer between HO3 of the reducing end residue and H2' (detected at C2') of the terminal residue. The occurrence of inter-residue hydrogen bonding, albeit transient, is judged important for the stabilization of three-dimensional structures, which may be essential in maintaining a conformational state for carbohydrate-protein interactions of glycans to take place in biologically important environments.

Water structuring above solutes with planar hydrophobic surfaces.

Many important biological solutes possess not only polar and hydrogen bonding functionalities, but also weakly-hydrating, or hydrophobic, surfaces. Theories of the hydration of such surfaces predict that their solvent interactions will change from a wetting type interaction to a dewetting regime as a function of the solute size, with a gradual transition in behavior taking place around characteristic lengths of ∼1 nm. Aggregations of non-polar species over this size range will undergo a transition from being dominated by entropy to being dominated by enthalpy. These transitions can be understood in part in terms of the geometries required of the solvating water molecules. We report here a series of simulations in aqueous solution of organic molecules with planar faces of increasing size, ranging from cyclopropane to circumcircumcoronene, in order to explore the transition in behavior for such solutes as their size increases. For this series, the dewetting transition occurred gradually, converging asymptotically to a limiting separation value for first layer water molecules of around 3.3 Å, while the transition in hydrogen bonding orientational structure occurred between cyclopropane and cyclopentadene. Water immediately adjacent to the largest planar hydrophobic surfaces oriented in ways that resembled on average the structural organization of the basal planes of ice.

Weakly hydrated surfaces and the binding interactions of small biological solutes.

Extended planar hydrophobic surfaces, such as are found in the side chains of the amino acids histidine, phenylalanine, tyrosine, and tryptophan, exhibit an affinity for the weakly hydrated faces of glucopyranose. In addition, molecular species such as these, including indole, caffeine, and imidazole, exhibit a weak tendency to pair together by hydrophobic stacking in aqueous solution. These interactions can be partially understood in terms of recent models for the hydration of extended hydrophobic faces and should provide insight into the architecture of sugar-binding sites in proteins.

Male-specific sesquiterpenes from Phyllotreta flea beetles.

Flea beetles in several genera are known to possess male-specific sesquiterpenes, at least some of which serve as aggregation pheromones that attract both sexes. In continuing research on the chemical ecology of Phyllotreta flea beetles, six new male-specific sesquiterpenes were identified, one from P. striolata (hydroxyketone 9) and five from P. pusilla (aldehydes 10-12 and 14 and alcohol 13); both species are crop pests. The minute amounts from beetles provided mass spectra and chromatographic data but were insufficient for complete structure determination. However, it was discovered that the new compounds could all be produced by applying organic reactions to previously identified flea beetle sesquiterpenes, and the resulting, larger amounts of material permitted definitive structure analysis by NMR. Molecular modeling was used in conjunction with NMR to define relative configurations of several newly created stereogenic centers. The absolute configurations of natural 9-14 were established by chiral gas chromatography/mass spectrometry. In electrophysiological tests (GC-EAD) conducted with P. striolata, compound 9 was detected with high sensitivity by the beetle antennae, which is consistent with a pheromonal function. The research opens new possibilities for using behavioral chemicals to monitor or manage these pest species.

Elucidation of the Crystal Growth Characteristics of SnO2 Nanoaggregates Formed by Sequential Low-Temperature Sol-Gel Reaction and Freeze Drying.

SnO2 nanoparticles are regarded as attractive, functional materials because of their versatile applications. SnO2 nanoaggregates with single-nanometer-scale lumpy surfaces provide opportunities to enhance hetero-material interfacial areas, leading to the performance improvement of materials and devices. For the first time, we demonstrate that SnO2 nanoaggregates with oxygen vacancies can be produced by a simple, low-temperature sol-gel approach combined with freeze-drying. We characterize the initiation of the low-temperature crystal growth of the obtained SnO2 nanoaggregates using high-resolution transmission electron microscopy (HRTEM). The results indicate that Sn (II) hydroxide precursors are converted into submicrometer-scale nanoaggregates consisting of uniform SnO2 spherical nanocrystals (2~5 nm in size). As the sol-gel reaction time increases, further crystallization is observed through the neighboring particles in a confined part of the aggregates, while the specific surface areas of the SnO2 samples increase concomitantly. In addition, X-ray photoelectron spectroscopy (XPS) measurements suggest that Sn (II) ions exist in the SnO2 samples when the reactions are stopped after a short time or when a relatively high concentration of Sn (II) is involved in the corresponding sol-gel reactions. Understanding this low-temperature growth of 3D SnO2 will provide new avenues for developing and producing high-performance, photofunctional nanomaterials via a cost-effective and scalable method.

27 ps DFT molecular dynamics simulation of alpha-maltose: A reduced basis set study.

DFT molecular dynamics simulations are time intensive when carried out on carbohydrates such as alpha-maltose. In a recent publication (Momany et al., J. Mol. Struct. THEOCHEM, submitted) forces for dynamics were generated from B3LYP/6-31+G* electronic structure calculations. The implicit solvent method COSMO was applied to simulate the solution environment. Here we present a modification of the DFT method that keeps the critical aspects of the larger basis set (B3LYP/6-31+G*) while allowing the less-essential atom interactions to be calculated using a smaller basis set, thus allowing for faster completion without sacrificing the interactions dictating the hydrogen bonding networks in alpha-maltose. In previous studies, the gg'-gg-c solvated form quickly converged to the "r" form during a 5 ps dynamics run. This important conformational transition is tested by carrying out a long 27 ps simulation. The trend for the "r" conformer to be most stable during dynamics when fully solvated, is confirmed, resulting in approximately 20/80% c/r population. Further, the study shows that considerable molecular end effects are important, the reducing end being fairly stable, the O6-H pointing at the O5, while the nonreducing end moves freely to take on different conformations. Some "kink" and transition state forms are populated during the simulation. The average H1'...H4 distance of 2.28 A confirms that the syn form is the primary glycosidic conformation, while the average C1'-O1'-C4 bond angle was 118.8 degrees , in excellent agreement with experimental values. The length of this simulation allowed the evaluation of vibrational frequencies by Fourier transform of the velocity correlation function, taken from different time segments along the simulation path.

Free energy surfaces for the interaction of D-glucose with planar aromatic groups in aqueous solution.

Multidimensional potentials of mean force for the interactions in aqueous solution of both anomers of D-glucopyranose with two planar aromatic molecules, indole and para-methyl-phenol, have been calculated using molecular dynamics simulations with umbrella sampling and were subsequently used to estimate binding free energies. Indole and para-methyl-phenol serve as models for the side chains of the amino acids tryptophan and tyrosine, respectively. In all cases, a weak affinity between the glucose molecules and the flat aromatic surfaces was found. The global minimum for these interactions was found to be for the case when the pseudoplanar face of ß-D-glucopyranose is stacked against the planar surfaces of the aromatic residues. The calculated binding free energies are in good agreement with both experiment and previous simulations. The multidimensional free energy maps suggest a mechanism that could lend kinetic stability to the complexes formed by sugars bound to sugar-binding proteins.

Ramachandran conformational energy maps for disaccharide linkages found in Burkholderia multivorans biofilm polysaccharides.

Ramachandran conformational energy maps have been prepared for all of the glycosidic linkages found in the C1576 exopolysaccharide that constitutes the biofilms of the bacterial species Burkholderia multivorans, a member of the Burkholderia cepacian complex that was isolated from a cystic fibrosis patient. This polysaccharide is a rhamnomannan with a tetrasaccharide repeat unit containing two mannose residues and two rhamnose residues, -[3-α-d-Man-(1→2)-α-d-Man-(1→2)-α-d-Rha-(1→3)-α-d-Rha-(1→]n-, where approximately 50% of the rhamnoses are randomly methylated on their O3 hydroxyl groups, further increasing the overall hydrophobicity of the chains. Because of the methylation, the tetrasaccharide repeat unit actually contains six possible linkages. The conformational energy maps are fully adiabatic relaxed maps in which the energy for each (ϕ,ψ) grid point on the map represents the lowest possible energy for the molecule in that conformation, considering all the combinations of the other degrees of freedom, such as hydroxyl orientations. Molecular dynamics simulations were used to verify that these maps indeed describe the conformational dynamics of these linkages. All six linkages were found to be quite restricted in possible ϕ angles, but to exhibit several possible low-energy ψ angles, suggesting that these chains could be quite flexible.

DFT conformational studies of alpha-maltotriose.

Recent DFT optimization studies on alpha-maltose improved our understanding of the preferred conformations of alpha-maltose. The present study extends these studies to alpha-maltotriose with three alpha-D-glucopyranose residues linked by two alpha-[1-->4] bridges, denoted herein as DP-3's. Combinations of gg, gt, and tg hydroxymethyl groups are included for both "c" and "r" hydroxyl rotamers. When the hydroxymethyl groups are for example, gg-gg-gg, and the hydroxyl groups are rotated from all clockwise, "c", to all counterclockwise, "r", the minimum energy positions of the bridging dihedral angles (phi(H) and psi(H)) move from the region of conformational space of (-, -), relative to (0 degrees , 0 degrees), to a new position defined by (+, +). Further, it was found previously that the relative energies of alpha-maltose gg-gg-c and "r" conformations were very close to one another; however, the DP-3's relative energies between hydroxyl "c" or "r" rotamers differ by more than one kcal/mol, in favor of the "c" form, even though the lowest energy DP-3 conformations have glycosidic dihedral angles similar to those found in the alpha-maltose study. Preliminary solvation studies using COSMO, a dielectric solvation method, point to important solvent contributions that reverse the energy profiles, showing an energy preference for the "r" forms. Only structures in which the rings are in the chair conformation are presented here.

Spectroscopy, dissociation dynamics, and potential energy surfaces for CN(A)-Ar.

The A (2)Pi-X (2)Sigma(+) band system of CN-Ar has been examined using fluorescence depletion and action spectroscopy techniques. Eight vibronic bands of the complex were observed in association with the monomer 3-0 transition. Pump-probe measurements were used to characterize CN(A (2)Pi(32),nu=3) fragments from direct photodissociation of CN(A (2)Pi,nu=3)-Ar and CN(X (2)Sigma(+),nu=7) fragments from CN(A (2)Pi,nu=3)-Ar predissociation. The latter showed a marked preference for population of positive parity diatomic rotational levels. Bound state calculations were used to assign the A-X bands and to obtain fitted potential energy surfaces for the A state. The average potential obtained from fitting had a well depth of D(e)=137.8 cm(-1). High-level ab initio calculations were used to obtain equilibrium Jacobi coordinates of theta(e)=94 degrees and R(e)=7.25 bohr. The near-symmetric character of the fitted potential energy surface was consistent with the symmetry preference observed in the predissociation dynamics.

Statistical Mechanics-Based Theoretical Investigation of Solvation Effects on Glucose Anomer Preferences.

The importance of solvation effects on the stability of glucose anomers has been studied by the combination of quantum mechanics and statistical mechanics, namely, the reference interaction site model self-consistent field spatial electron density distribution. The preferences of α- and ß-glucose in H2O are well reproduced with the obtained ratio of 35:65 for α- and ß-glucose, respectively. Indirect interactions and bulk effects, described by the Onsager model, are relatively small compared to the direct solute-solvent interactions, especially in [DMIM]Cl and dimethyl sulfoxide. From the decomposition of solvation free energy and solvation structures, it can be seen that the interactions with the solvent molecules greatly contribute to the anomer preferences.

Performance of Density-Functional Tight-Binding in Comparison to Ab Initio and First-Principles Methods for Isomer Geometries and Energies of Glucose Epimers in Vacuo and Solution.

Density functional theory (DFT) is a widely used methodology for the computation of molecular and electronic structure, and we confirm that B3LYP and the high-level ab initio G3B3 method are in excellent agreement for the lowest-energy isomers of the 16 glucose epimers. Density-functional tight-binding (DFTB) is an approximate version of DFT with typically comparable accuracy that is 2 to 3 orders of magnitude faster, therefore generally very suitable for processing large numbers of complex structures. Conformational isomerism in sugars is well known to give rise to a large number of isomer structures. On the basis of a comprehensive study of glucose epimers in vacuo and aqueous solution, we found that the performance of DFTB is on par to B3LYP in terms of geometrical parameters excluding hydrogen bonds and isomer energies. However, DFTB underestimates both hydrogen bonding interactions as well as torsional barriers associated with rotations of the hydroxy groups, resulting in a counterintuitive overemphasis of hydrogen bonding in both gas phase as well as in water. Although the associated root mean squared deviation from B3LYP within epimer isomer groups is only on the order of 1 kcal/mol, this deviation affects the correct assignment of major isomer ordering, which span less than 10 kcal/mol. Both second- as well as third-order DFTB methods are exhibiting similar deviations from B3LYP. Even after the inclusion of empirical dispersion corrections in vacuum, these deviations remain for a large majority of isomer energies and geometries when compared to dispersion-corrected B3LYP.

DFT studies of the disaccharide, alpha-maltose: relaxed isopotential maps.

The disaccharide, alpha-maltose, forms the molecular basis for the analysis of the structure of starch, and determining the conformational energy landscape as the molecule oscillates around the glycosidic bonds is of importance. Thus, it is of interest to determine, using density functionals and a medium size basis set, a relaxed isopotential contour map plotted as a function of the phi(H) and psi(H) dihedral angles. The technical aspects include the method of choosing the starting conformations, the choice of scanning step size, the method of constraining the specific dihedral angles, and the fitting of data to obtain well defined contour maps. Maps were calculated at the B3LYP/6-31+G( *) level of theory in 5 degrees intervals around the (phi(H),psi(H))=(0 degrees ,0 degrees ) position, out to approximately +/-30 degrees or greater, for gg-gg'-c, gg-gg'-r, gt-gt'-c, gt-gt'-r, tg-tg'-c, and tg-tg'-r conformers, as well as one-split gg(c)-gg'(r) conformer. The results show that the preferred conformation of alpha-maltose in vacuo depends strongly upon the hydroxyl group orientations ('c'/'r'), but the energy landscape moving away from the minimum-energy position is generally shallow and transitions between conformational positions can occur without the addition of significant energy. Mapped deviations of selected parameters such as the dipole moment; the C1-O1-C4', H1-C1-O1, and H4'-C4'-O1 bond angles; and deviations in hydroxymethyl rotamers, O5-C5-C6-O6, O5'-C5'-C6'-O6', C5-C6-O6-H, and C5'-C6'-O6'-H', are presented. These allow visualization of the structural and energetic changes that occur upon rotation about the glycosidic bonds. Interactions across the bridge are visualized by deviations in H(O2)...O3', H(O3')...O2, and H1...H4' distances and the H(O2)-O2-C2-C1 and H'(O3')-O3'-C3'-C4' hydroxyl dihedral angles.

DFT study of alpha- and beta-D-allopyranose at the B3LYP/6-311++G ** level of theory.

One hundred and two conformations of alpha- and beta-D-allopyranose, the C-3 substituted epimer of glucopyranose, were geometry optimized using the density functional, B3LYP, and the basis set, 6-311++G **. Full geometry optimization was performed on different ring geometries and on the hydroxymethyl rotamers (gg/gt/tg). Analytically derived Hessians were used to calculate zero point energy, enthalpy, and entropy. The lowest energy and free energy conformation found is the alpha-tg(g-)-4C1-c conformation, which is only slightly higher in electronic (approximately 0.2 kcal/mol) and free energy than the lowest energy alpha-D-glucopyranose. The in vacuo calculations showed a small (approximately 0.3 kcal/mol) energetic preference for the alpha- over the beta-anomer for allopyranose in the 4C1 conformation, whereas in the 1C4 conformation a considerable (approximately 1.6 kcal/mol) energetic preference for the beta- over the alpha-anomer for allopyranose was encountered. The results are compared to previous aldohexose calculations in vacuo. Boat and skew forms were found that remained stable upon gradient optimization although many starting boat conformations moved to other skew forms upon optimization. As found for glucose, mannose, and galactose the orientation and interaction of the hydroxyl groups make the most significant contributions to the conformation/energy relationship in vacuo. A comparison of different basis sets and density functionals is made in the Discussion section, confirming the appropriateness of the level of theory used here.

DFT study of alpha- and beta-D-galactopyranose at the B3LYP/6-311++G** level of theory.

Forty-one conformations of alpha- and beta-d-galactopyranose were geometry optimized using the B3LYP density functional and 6-311++G** basis set. Full geometry optimization was performed on different ring geometries and different hydroxymethyl rotamers (gg/gt/tg). Analytically derived Hessians were used to calculate zero point energy, enthalpy, and entropy. The lowest energy and free-energy conformation found is the alpha-gg-(4)C(1)-c chair conformation, which is of lower electronic and free energy than the lowest energy alpha-d-glucopyranose conformer because of favorable hydrogen-bonding interactions. The in vacuo calculations showed considerable ( approximately 2.2kcal/mol) energetic preference for the alpha over the beta anomer for galactopyranose in both the (4)C(1) and (1)C(4) chair conformations. Results are compared to glucopyranose and mannopyranose calculations in vacuo. Boat and skew-boat forms were found that remained stable upon gradient optimization, although many starting conformations moved to other boat forms upon optimization. As with glucopyranose and mannopyranose, the orientation and interaction of the hydroxyl groups make the most significant contributions to the conformation-energy relationship in vacuo.

Comparison of the simulations of cellulosic crystals with three carbohydrate force fields.

Three independently developed molecular mechanics force fields for carbohydrates have been used to simulate a suite of small molecule analogs of cellulose for which crystal structures have been reported, as a test to determine which might be best for simulations of cellulose itself. Such evaluation is necessary since the reported cellulose crystal structure is not stable in molecular dynamics simulations with any available force field. The present simulations found that all three resulted in small deviations from the reported crystal structures, but that all were reasonably accurate and none was clearly superior to the others for the entire suite of structures examined.

Water structuring over the hydrophobic surface of cellulose.

Many important biological solutes possess not only polar and hydrogen-bonding functionalities but also weakly hydrating, or hydrophobic, surfaces. While the aggregation of these hydrophobic surfaces has been shown to play an important role in the aggregation of individual chains of cellulose, it is not known whether the water structuring imposed by these hydrophobic surfaces more closely resembles that associated with small hydrophobic solutes like methane and fats or more closely resembles that associated with extended hydrophobic surfaces like mica or waxy planes. By using molecular dynamics simulations to characterize the water molecule orientations over different regions of the 100 surface of cellulose in contact with water, it was found that the hydrophobic strips of the cellulose crystal are sufficiently narrow that they hydrate like a fatty acid chain, rather than like a more extended surface, suggesting that their aggregation would be dominated by entropy rather than enthalpy.

DFTr studies of five- and six-residue cyclic-ß(1→4) cellulosic molecules.

Density functional (DFT) conformational in vacuo studies of cellobiose have shown that ϕ(H) -anti conformations are low in energy relative to the syn forms, while the ψ(H) -anti forms are higher in energy. Further, as the cellulosic fragments became larger than a disaccharide and new hydrogen bonding interactions between multiple residues become available, stable low energy ϕ(H) -anti, and ψ(H) -anti cellulosic structures became possible. To test the stability of cyclic anti-conformations, a number of ß-linked five- and six-residue molecules were created and then energy optimized in solvent (water, n-heptane) using the implicit solvation method COSMO at the B3LYP level of theory. The created symmetric cyclic structures were without distortion. Upon optimization some cyclic conformations were found to be of low energy when compared with linear five- and six-residue chains, after correcting the energy for the exclusion of a water molecule upon cyclization. It was also obvious from the hydrogen bonding network formed above and below the plane of the cyclic structure that these structures could exhibit strong synergistic tendencies. The conformational energy preferences for clockwise "c" and counter-clockwise "r" hydroxyl groups and preference for the hydroxymethyl rotamers is described. Because these structures contain energetically unfavorable flipped conformations in water, that is, dihedral angles of ∼180°/0° or ∼0°/180° in ϕ(H) /ψ(H) , it is clear that the synthesis of these compounds will be challenging.

DFT energy optimization of a large carbohydrate: cyclomaltohexaicosaose (CA-26).

CA-26 is the largest cyclodextrin (546 atoms) for which refined X-ray structural data is available. Because of its size, 26 D-glucose residues, it is beyond the scope of study of most ab initio or density functional methods and to date has only been computationally examined using empirical force fields. The crystal structure of CA-26 is folded like a figure "8" into two 10 D-glucoses long antiparallel left-handed V (Verkleisterung)-type helices with a "band-flip" and "kink" at the top and bottom of the helices. DFTr methods were applied to CA-26 to determine if a carbohydrate molecule of this size could be geometry optimized and if it would show structural variances from application of dispersion and/or solvation. The DFTr reduced basis set method developed by the authors uses 4-31G on the carbon atoms of the glucose rings and 6-31+G* on all other atoms. B3LYP is the density functional used to successfully optimize CA-26, and other density functionals were then applied, including a self-consistent charge density functional tight binding (SCC-DFTB) method and the B97D (dispersion-corrected) and B97D-PCM (dispersion + implicit solvent) methods. Heavy atom coordinates were taken from one X-ray structure, fitted with hydrogen atoms, and geometry optimized using PM3 followed by B3LYP/6-31+G*/4-31G optimization. After optimization, the heavy atom rms deviation of the optimized DFTr (B3LYP) structure to the crystal structure was 0.89 Å, the rmsd of the B97D optimization was 1.38 Å, that for B97D-PCM was 0.95 Å, and that for SCC-DFTB was 0.94 Å. These results are very good considering that no explicit water molecules were included in the computational analysis and there were ~32-38 water molecules around each CA-26 molecule in the crystal structure. Tables of internal coordinates and puckering parameters were compared to the X-ray structures, and close correspondence was found.

Caffeine and sugars interact in aqueous solutions: a simulation and NMR study.

Molecular dynamics simulations were carried out on several systems of caffeine interacting with simple sugars. These included a single caffeine molecule in a 3 m solution of α-D-glucopyranose, at a caffeine concentration of 0.083 m, a single caffeine in a 3 m solution of ß-D-glucopyranose, and a single caffeine molecule in a 1.08 m solution of sucrose (table sugar). Parallel nuclear magnetic resonance titration experiments were carried out on the same solutions under similar conditions. Consistent with previous thermodynamic experiments, the sugars were found to have an affinity for the caffeine molecules in both the simulations and experiments, and the binding in these complexes occurs by face-to-face stacking of the hydrophobic triad of protons of the pyranose rings against the caffeine face, rather than by hydrogen bonding. For the disaccharide, the binding occurs via stacking of the glucose ring against the caffeine, with a lesser affinity for the fructose observed. These findings are consistent with the association being driven by hydrophobic hydration and are similar to the previously observed binding of glucose rings to various other planar molecules, including indole, serotonin, and phenol.